Plasmodium falciparum is the causative agent responsible for the most severe form of human malaria, a disease that kills more than a million people a year, mostly young children in Africa. These protozoan parasites invade and ultimately destroy circulating red blood cells (RBCs) of their host, leading to severe anemia and the frequently lethal syndromes of cerebral malaria and pregnancy associated malaria. Over the course of an infection, small sub-populations of parasites arise that have an altered antigenic phenotype, thus avoiding the antibody response of the host. This process is referred to as antigenic variation and is responsible for the persistent nature of the disease as well as the waves of parasitemia frequently observed in P. falciparum infections. Antigenic variation of P. falciparum infected RBCs results from switches in expression between individual members of the multi-copy var gene family. Each var gene encodes a different form of a protein called PfEMP1. This protein is placed on the surface of the infected RBCs and mediates adhesion to specific receptors found on the endothelial surfaces of the blood vessel walls of the infected individual. This adhesion is responsible for many of the disease manifestations of infection with P. falciparum, including both cerebral malaria and pregnancy associated malaria. Only a single var gene is expressed at a time by any given parasite, thus determining both the antigenic phenotype of the infected cells as well as their adhesive properties. Therefore var gene expression is the heart of both antigenic variation and virulence of malaria infections. This project is designed to understand the molecular mechanisms the regulate var gene expression and antigenic variation by malaria parasites. Previous work has demonstrated that mutually exclusive expression of var genes involves specific modifications to the chromatin structure at each var gene, however how these modifications are coordinated to result in expression of only a single gene at a time out of a family of 60 remains unknown. The specific aims of the project are 1) to identify and characterize chromatin boundary/insulator elements that regulate chromatin assembly at var genes, 2) to determine the role of noncoding RNAs in targeting chromatin assembly to var gene loci, and 3) to investigate the mechanism of translational regulation of a specific var gene implicated in pregnancy associated malaria. The experimental design relies on creating transgenic parasite lines using reporter genes and drug selectable markers to rapidly determine the role of these regulatory elements in controlling var gene expression. The long-term goals are to develop methods to disrupt the process of antigenic variation and thereby shorten the length of an infection and reduce its severity.